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An increase in intracellular cyclic AMP modulates nitric oxide production in IFN-gamma-treated macrophages
D Mullet1, R H Fertel, D Kniss
1Department of Pharmacology, Ohio State University College of Medicine, Columbus 43210, USA.
Abstract:
Macrophages treated with IFN-gamma alone are stimulated to produce nitric oxide. The level of nitric oxide production can be enhanced significantly when IFN-gamma treatment is combined with other agents (e.g., LPS, TNF-alpha, IL-2, etc.). We tested the hypothesis that cAMP plays a role in the IFN-gamma-induced activation of macrophages. Our experiments indicate that factors that increase the concentration of cAMP in the murine macrophage cell line ANA-1 can also enhance IFN-gamma-induced production of nitric oxide. PGE2 and cholera toxin increased the production of nitrite (an indicator of nitric oxide production) in IFN-gamma-treated ANA-1 macrophages by at least twofold. These factors produced no increase in nitric oxide production in the absence of IFN-gamma treatment. The increase in nitric oxide production corresponded to an increase in the accumulation of nitric oxide synthase mRNA without a change in stability of mRNA. Dibutyryl cAMP and Sp-cAMPs (a selective activator of cAMP-dependent protein kinase I and II) also increased nitric oxide production in IFN-gamma-treated macrophages. However, at very high concentrations (i.e., >100 microM), the stimulatory effect was decreased. These studies indicate that elevation of intracellular cAMP causes a dose-dependent, biphasic alteration of IFN-gamma-induced nitric oxide production in murine macrophages. Moreover, they suggest that agents that affect nitric oxide synthesis may do so via modulation of the cAMP second messenger system.
Insights
Cyclic AMP (cAMP) enhances nitric oxide production in macrophages activated by interferon-gamma (IFN-gamma). This suggests cAMP signaling pathways modulate macrophage immune responses and nitric oxide synthesis.
Area of Science:
- Immunology
- Cellular Biology
- Biochemistry
Background:
- Macrophages are key immune cells involved in host defense.
- Interferon-gamma (IFN-gamma) activates macrophages to produce nitric oxide (NO).
- NO production by macrophages can be modulated by various signaling molecules.
Purpose of the Study:
- To investigate the role of cyclic adenosine monophosphate (cAMP) in IFN-gamma-induced nitric oxide production in macrophages.
- To determine if elevated cAMP levels enhance NO synthesis in response to IFN-gamma.
Main Methods:
- Murine macrophage cell line (ANA-1) was treated with IFN-gamma.
- Agents known to increase intracellular cAMP (PGE2, cholera toxin, dibutyryl cAMP, Sp-cAMPs) were added.
- Nitrite production (an indicator of NO) and nitric oxide synthase (NOS) mRNA levels were measured.
- The effect of varying cAMP concentrations was assessed.
Main Results:
- Factors increasing cAMP significantly enhanced IFN-gamma-induced nitrite production (at least twofold).
- These cAMP-elevating agents had no effect on NO production without IFN-gamma.
- Increased NO production correlated with increased NOS mRNA accumulation, not mRNA stability.
- A dose-dependent, biphasic effect of cAMP on NO production was observed, with inhibition at high concentrations (>100 microM).
Conclusions:
- Intracellular cAMP plays a significant role in modulating IFN-gamma-induced nitric oxide production in murine macrophages.
- Elevating cAMP levels can enhance NO synthesis, suggesting a mechanism for immune response modulation.
- These findings indicate that the cAMP second messenger system is involved in regulating nitric oxide synthesis in macrophages.